The Melbourne Rebels have missed out on their first finals appearance after the Sharks beat the Jaguares in Durban to secure eighth position and the last quarter-final slot.
And in further bad news for Australian teams from South Africa, the Waratahs slipped down to third spot on the ladder after the Lions beat the Bulls.
Instead of hosting the Jaguares, the Waratahs will host the Highlanders in Sydney on Saturday night.
The Rebels' narrow loss to the Highlanders in Dunedin on Saturday meant they were banking on the Jaguares to continue on their winning run at Kings Park in the last regular season game.
But in a hard-fought clash, the South African side ground out a 20-10 win over the Jaguares and finished level on 36 points with the Rebels.
The Sharks also finished the season with superior for-and-against, however, so they stayed above the Rebels and locked down eighth spot. The difference was just 16 points.
It was a heartbreaking finish to the season for the Rebels, who led the Highlanders inside the final quarter but conceded three tries to lose.
They also blew their chance to pick up a vital win with a mistake-riddled loss to Queensland a week earlier.
The Waratahs will also be kicking themselves after letting second place slip from their grasp, and a harder route to any potential Super Rugby victory.
The NSW side crashed to a sleepy defeat to the Brumbies at home, and then watched as the Lions secured top spot in the African conference with a win over the Bulls in Johannesburg.
That result gave the Lions second spot, and a quarter-final against the Jaguares.
The Waratahs finished third and will play the Highlanders, who they beat earlier this season at Allianz Stadium but after the Kiwi side were reduced to 14 men.
Even if they beat the Highlanders, the Waratahs would then have to travel to Johannesburg if the Lions beat the Argentinian side next weekend.
They'll be hoping they can beat the Highlanders and, yet again, be banking on the Jaguares to pull off an upset win over the Lions, so they can host a semi-final a week later.
2018 Super Rugby quarter-finals
QF4: Hurricanes (4th) v Chiefs (5th), Friday July 20, 5:35pm AEST
QF1: Crusaders (1st) v Sharks (8th), Saturday July 21, AMI Stadium, 5:35pm AEST
QF3: Waratahs (3rd) v Highlanders (6th), Saturday July 21, Allianz Stadium, 8:05pm AEST
QF2: Lions (2nd) v Jaguares (7th), Sunday July 22, Ellis Park, 1:05am AEST